Androgenesis of Red Cabbage in Isolated Microspore Culture In Vitro.

Red cabbage belongs to the economically important group of vegetable crops of the Brassicaceae family. A unique feature of this vegetable crop that distinguishes it from other members of the family is its unique biochemical composition characterized by high anthocyanin content, which gives it antioxidant properties. The production mainly uses F1 hybrids, which require constant parental lines, requiring 6-7 generations of inbreeding. Culture of isolated microspores in vitro is currently one of the promising methods for the accelerated production of pure lines with 100% homozygosity. The aim of this study is to investigate the factors and select optimal parameters for successful induction of red cabbage embryogenesis in isolated microspore culture in vitro and subsequent regeneration of DH plants. As a result of research, for the first time, it was possible to carry out the full cycle of obtaining DH plants of red cabbage from the induction of embryogenesis to their inclusion in the breeding process. The size of buds containing predominantly microspores at the late vacuolated stage and pollen at the early bi-cellular stage has to be selected individually for each genotype, because the embryoid yield will be determined by the interaction of these two factors. In the six samples studied, the maximum embryoid yield was obtained from buds 4.1-4.4 mm and 4.5-5.0 mm long, depending on the genotype. Cultivation of microspores was carried out on liquid NLN culture medium with 13% sucrose. The maximum number of embryoids (173.5 ± 7.5 pcs./Petri dish) was obtained on culture medium with pH 5.8 and heat shock at 32 °C for 48 h. Successful embryoid development and plant regeneration by direct germination from shoot apical meristem were achieved on MS culture medium with 2% sucrose and 0.7% agar, supplemented with 6-benzylaminopurine at a concentration of 1 mg/L. Analysis of the obtained regenerated plants, which successfully passed the stage of adaptation to ex vitro conditions by flow cytometry, showed that most of them were doubled haploids (up to 90.9%). A low number of seeds produced by self-fertilization in DH plants was observed.

Production of Gynogenic Plants of Red Beet (Beta vulgaris L.) in Unpollinated Ovule Culture In Vitro.

The unique and balanced components of the biochemical composition, together with high antioxidant activity, make the red beet necessary a dietary vegetable crop, much contributing to healthy food ration. The application of the technology for producing gynogenic plants in vitro increases the genetic diversity and significantly reduces the period of time required to obtain the appropriate homozygous lines used to create the F1 hybrids that are demanded in the market. For induction of gynogenesis, we used IMB medium developed by us with the addition of 55 g/L sucrose, 3 g/L phytogel, 200 mg/L ampicillin, and 0.4 mg/L thidiazuron (TDZ) and cultured at 28 °C in the dark for 4-6 weeks. Shoot regeneration from embryoids and callus was performed on MS medium with 20 g/L sucrose, 3 g/L phytogel, 1 mg/L 6-benzylaminopurine (BAP), and 0.1 mg/L gibberellic acid (GA3). Immersion of the obtained microshoots with 5-7 well-developed leaves for 10-15 s into concentrated sterile indole-3-butyric acid (IBA) solution (50 mg/L) followed by their cultivation on solid medium ½ IMB with 2% sucrose and 3 g/L phytogel was the most efficient method for root formation. The addition of silver nitrate (22 mg/L) to the nutrient medium provoked an increase in the number of induced ovules up to nine per Petri dish (up to 25% of induced ovules). Gynogenic development was produced in six out of 11 genotypes studied, and the plants that were then acclimatized to ex vitro conditions were obtained in three genotypes (Nezhnost', Dobrynya, b/a 128). The evaluation of ploidy of gynogenic plants that was carried out by flow cytometry and direct counting of chromosomes stained with propion-lacmoide revealed that all obtained gynogenic plants were haploids (2n = x = 9).

Effects of Genotype and Culture Conditions on Microspore Embryogenesis and Plant Regeneration in Brassica Rapa ssp. Rapa L.

Turnip is a biennial crop and, consequently, the creation of pure lines for breeding is a time-consuming process. The production of pure turnip lines using doubled haploids produced in isolated microspore culture has not been sufficiently developed. The aim of the present work was to determine some key factors inducing embryogenesis in the isolated microspore culture of turnip, as well as investigating the manners of embryo development. It was shown that the acidity of the medium is an important factor in embryo production; different optimal pH levels ranging from 6.2 to 6.6 corresponded to individual genotypes. Such factors as the cold treatment of buds and the addition of activated charcoal to the nutrient medium increased the responsiveness of all genotypes studied. The turnip variety 'Ronde witte roodkop herfst' demonstrated a genetic disorder in the development of microspores; namely, non-separation of some microspores from tetrads. In the in vitro culture, each of the daughter microspores developed on its own. This indicates the dependence of the possibility of embryogenesis in the turnip microspore culture on the genotype. Results suggest that the initiation of secondary embryogenesis in primary embryos leads to an increase in the proportion of doubled haploid plants.

Effect of Beta-Lactam Antibiotics on Microspore Embryogenesis in Brassica Species.

Antibiotics are widely applied for plant cultivation in vitro to eliminate bacterial contamination. However, they can have both positive and negative effects on the cells of cultivated plants, and these effects largely depend on the type antibiotic used and its concentration. The objective of the present study was to estimate the effect of ß-lactam antibiotics ampicillin (Amp) and cefotaxime (Cef) on microspore embryogenesis induction in vitro in the Brassica species. The performed experiments confirmed cefotaxime inhibits microspores in B. napus and B. oleracea, even in concentrations as low as 50 mg/L. The highest embryo yield was obtained for B. napus in the NLN-13 medium with added ampicillin in concentrations of 50-100 mg/L as an antimicrobial agent. This embryo yield was significantly higher than that obtained in a medium without supplemented antibiotics and two times higher than in the medium with added cefotaxime. Analogous results were obtained for B. oleracea and B. rapa.

Yield, Growth, Quality, Biochemical Characteristics and Elemental Composition of Plant Parts of Celery Leafy, Stalk and Root Types Grown in the Northern Hemisphere.

Celery is one of the major nutraceutical vegetable species due to the high dietary and medicinal properties of all of its plant parts. Yield, growth and produce quality of six celery genotypes belonging to leafy (Elixir and Samurai), stalk (Atlant and Primus) or root (Egor and Dobrynya) types, as well as the distribution of biomass, sugars, mineral elements and antioxidants among the different plant parts, were assessed. Within the celery root type, cultivar Dobrynya resulted in higher yield than Egor, whereas the genotype did not significantly affect the marketable plant part production of leafy and stalk types. Leaf/petiole ratios relevant to biomass, total dissolved solids, sugars, ascorbic acid, flavonoids, antioxidant activity and ash, K, Zn, Fe, Mn, Cu and Se content were significantly affected by the celery type examined. Ash content was highest in the leaves and lowest in the roots. Celery antioxidant system was characterized by highly significant relationships between ascorbic acid, polyphenols, flavonoids, antioxidant activity and Zn. Among the celery types analyzed, the highest values of chlorophyll, Fe and Mn content as well as antioxidant activity were recorded in leaves from root genotypes, which suggests interesting nutraceutical prospects of the aforementioned plant parts for human utilization.